Abstract

Membranous fat necrosis (MFN) in the gallbladder is rare. We found marked MFN as the cause of peculiar foreign body reaction in a Japanese woman in her late seventies, with severe chronic cholecystitis. Histopathological examination of the extirpated gallbladder revealed severe grade chronic cholecystitis accompanied by many multinucleated giant cells and phagocytizing yellowish wavy membranous substance in the subserosal layer. The results of Sudan black B, Luxol fast blue, and Ziehl-Neelsen staining of the substance were all positive. CD68 antibody was strongly positive in multinucleated giant cells. These findings suggested that the membranous structures were the products of MFN, which was recognized as a foreign body and phagocytized by macrophages. Stenosis of intramural sclerotic arteries induced chronic circulatory disturbances leading to MFN that resulted in a peculiar foreign body reaction in the subserosal fatty tissues of the gallbladder. To our best knowledge, these findings have not been reported in the literature thus far.

Keywords

Introduction

Membranous fat necrosis (MFN) [1-5], membranocystic
change [1,6,7], or lipomembranous panniculitis [8,9] are
usually found in skin-related diseases[7,9,10,11] and rarely
occur in organs other than the skin [2-5]. However, if
these changes are systemic, the brain and skeletal systems
are mainly affected. Nasu et al. reported such a case for
the first time in 1973 [6]. Subsequently, focal MFN was
accidentally detected as one of the main histopathological
changes in various kinds of skin-related diseases, such as
systemic lupus erythematosus [7,9,11], systemic sclerosis
[10], diabetes [1,8], and scrotal and/or perineal lipogranuloma [12,13]. MFN has rarely been reported in the breast
[2], lipoma [3], appendices epiploicae[4], and mature
teratoma of the ovary [5].

In this study, we present a case of MFN in the gallbladder,
which is extremely rare. We detected MFN in the extirpated
gallbladder in only 1 case among 1,538 consecutive
cases during the past 13.5 years in Matsuyama-shimin
Hospital. Interestingly, a marked foreign body reaction
was detected, accompanied by multinucleated giant cells
and phagocytizing peculiar membranous structures of MFN. We observed that phagocytized MFN was stained
distinctly with Sudan black B (SBB) in a dewaxed section.

Case report

The patient was a Japanese woman in her late seventies
who had undergone laparoscopic resection of the gallbladder.
The wall of gallbladder was thickened due to
chronic inflammation caused by hemorrhages and severe
fibrosis (Fig. 1). The extirpated gallbladder was fixed in
10% formalin solution and embedded in paraffin. The
dewaxed sections were stained with hematoxylin-eosin
(HE), periodic acid-Schiff (PAS), and diastase-digested
PAS (D-PAS), elastica van Gieson, Berlin blue, Victoria
blue, orcein, Schmorl, prolonged Ziehl-Neelsen (Z-N),
SBB, and Luxol fast blue (LFB) stains. Immunohistochemical
staining was performed, as described previously
[13,14], by using labeled streptavidin-biotin (LSAB)2
kit/horseradish peroxidase (HRP) (DakoCytomation,
Kyoto, Japan) with diaminobenzidine as the substrate and
antibodies against macrophage-associated antigen CD68
(1:50, pronase pretreated; Dako, Kyoto, Japan) and S-100
protein (1:100, no pretreatment; Dako, Kyoto, Japan).

A written informed consent has been obtained from the
patient, and her identity has been protected.

Pathology

The entire fundamental mucosa was not retained and its
surface was widely denuded in the extirpated gallbladder
(Fig. 1). No epithelial covering cells were found on the
surface. The wall was thickened due to fibrosis and severe
hemorrhage, revealing several cystic sinuses and many
superficial macrophages containing hemosiderin. In the
subserosal layer, peculiar multinucleated giant cells were
scattered (Fig. 2a) and gathered focally, and these cells
contained light yellowish membranous structures in their
cytoplasm (Fig. 2b). A membranous yellowish substance
was also embedded in the cell wall without an accompanying
foreign body reaction. Fine membranous products
of MFN were obviously phagocytized by macrophages
(Fig. 2b). These findings were often detected in the indicated
areas in Fig. 1. D-PAS reaction was positive for
phagocytized MFN. In order to identify these structures
precisely, SSB staining was performed using dewaxed sections; this staining revealed distinct peculiar black
membranous structures in the cytoplasm of multinucleated
giant cells (Fig. 3a). The surface of MFN was fluffy
and had a microwave-like pattern (Fig. 3b). LFB also
showed a complicated lamellar structure in the cytoplasm
of multinucleated giant cells (Fig. 4). The cytoplasm of
multinucleated giant cells was positive (Fig. 5), but the
phagocytized membranous substance was negative for
CD68 antibody. S100 protein antibody was negative for
all the giant cells. Prolonged Z-N staining also gave positive
results for MFN. However, the results of Berlin blue,
orcein, Schmorl, Victoria blue, and elastica van Gieson
staining were all negative. Thus, the results of SBB as
well as prolonged Z-N, D-PAS(focal), and LFB staining
were all positive for MFN in dewaxed sections. These
results suggested that complicated membranous structures
in the cytoplasm of multinucleated giant cells were produced
by MFN.

With regard to the histogenesis of MFN in this case, MFN
was mostly thought to be caused by chronic circulatory
disturbances. The stenotic or obstructed arteries due to
arteriosclerosis were often detected in the wall (Fig. 6a),
which was clearly visible after elastic fiber staining(Fig.
6b&c). Further, marked fibrosis, hemorrhage, and inflammatory
cell infiltration were also observed in the wall,
which appeared to be other causes of ischemic reaction in
the gallbladder.

Discussion

MFN was usually found in the skin-related diseases
[7,9,10,12] in patients with obesity [1], diabetes [1,8], or
circulatory disturbance [15], but it was rarely found in the
breast [2], lipoma [3], appendices epiploicae [4], and mature
cystic teratoma of ovary [5]. MFN of the gallbladder
has not yet been reported in the literature, though various
kinds of chronic inflammation are extremely common in
this organ.

In the present study, the pathological examination of a
patient with marked chronic cholecystitis revealed peculiar
MFN in subserosal fatty tissues of its wall accompanied
by severe fibrosis and hemorrhage. Further, many
macrophages and phagocytized MFN products were detected
in the subserosal area. Generally, peripheral circulatory
disturbances are thought to be the cause of MFN
[1,3,4,8,9,15]. Although our patient did not have either
diabetes or cutaneous lesions of collagen vascular diseases,
we suspected some kinds of circulatory disturbance
in its wall due to arteriosclerotic changes associated with
marked fibrosis and hemorrhage. The involved fatty tissues
became ischemic and induced MFN. These degenerated
fatty tissues were recognized as foreign body because
there was a marked change in the tissue characteristics
due to ischemia. MFN induced macrophage aggregation,
and it was subsequently phagocytized into the cytoplasm
and retained in multinucleated giant cells as a foreign
body. These phagocytized MFN might be digested in
different ways, depending on each MFN.

With regard to the staining characteristics of MFN in the
present case, the results of D-PAS [3,13], SBB [3,10,12],
LFB [10], and prolonged Z-N [3,4] staining were positive
for MFN, and immunohistochemistry of CD68 was clearly
positive for multinucleated giant cells [13]. The results
of the present case suggested that MFN was not like a
ceroid, as reported previously [3,4,5], because it gave
negative results with orcein, Victoria blue, and Schmorl
stains. MFN might be heterogeneous because the nature
of degeneration or denaturation might differ in each case.
However, MFN was positive at least for SBB, LFB, and
prolonged Z-N as observed in our case.

In keeping with these findings, the real cause of phagocytizing
is unclear. Phagocytosis of a similar nature has
been observed in sclerosing lipogranuloma of scrotum or
perineal region [12,13], and in these cases, MFN was
phagocytized by macrophages like in our case. However,
MFN was not phagocytized in some cases [8]. Thus far, to
our best knowledge, there has been no report on MFN of
gallbladder in the literature. Further investigation is needed
to identify the precise reasons of the rarity of MFN in
gallbladder.

Acknowledgment

The authors are grateful to Mrs. Y. Matsuka, Ms. W. Tanihata,
and Mrs. M. Izumimoto for their technical assistance
and to Ms. K. Matsushita and Ms. K. Takasuka for their
secretarial assistance.